Toxoplasma gondii is a common parasitic infection of humans where it normally causes uncomplicated yet persistent infections. Reactivation of chronic infection can lead to severe disease in immunocompromised patients, including those with HIV-AIDS. With the advent of HAART therapy, the frequency of toxoplasmosis has decreased. However, there are still many areas of the world, and susceptible patient populations, were advanced HAART therapy is not widely or effectively implemented. Hence, toxoplasmosis presents a continued risk to immunocompromised patients due to the high prevalence of chronic infection and limitations of existing antibiotics, which do not eradicate chronic infections. Our studies are designed to define the molecular basis of pathogenesis in T. gondii, focusing on more virulent parasite lineages that may pose greater risk in immunocompromised patients. We have previously used genetic analyses of the clonal lineages of North America to identify virulence genes in T. gondii. However, T. gondii strains are much more diverse and undergo greater genetic exchange in other regions of the world. For example, the majority of South American strains are acutely virulent in the mouse model and such strains are frequently found in HIV positive patients where they cause severe CNS disease. The basis for this enhanced pathogenicity is presently unknown, but importantly, it cannot be explained by known virulence determinants. In preliminary studies, we have developed several new genetic crosses between highly virulent South American strains and less virulent lineages typical of North America. The proposed studies will use genetic mapping to identify virulence determinants of T. gondii and test the role of these parasite effectors in mediating survival in human cells. In preliminary studies, we have shown that IFN-?-activated human cells restrict the growth of T. gondii in a strain-specific manner. The proposed studies will define the cellular and molecular basis of this resistance pathway in human cells. Genetic approaches will be used to identify the molecular determinants of parasite virulence. The overall goal of our studies is to define the molecular mechanisms by which T. gondii overcomes host resistance, thus enhancing its survival and leading to more severe disease. Successful identification of parasite virulence factors may lead to improved detection and/or improved interventions designed to combat infection.